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| Status |
Public on Aug 05, 2021 |
| Title |
CLAMP_MTD_0-2_hr_1 |
| Sample type |
SRA |
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| Source name |
embryos 0-2hr
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| Organism |
Drosophila melanogaster |
| Characteristics |
developmental stage: embryos 0-2hr
genotype: MTD
chip antibody: CLAMP
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| Growth protocol |
To deplete maternally deposited clamp or zld mRNA throughout oogenesis, we crossed a maternal triple driver (MTD-GAL4, Bloomington, #31777) line with a Transgenic RNAi Project (TRiP) clamp RNAi line (Bloomington, #57008) or a TRiP zld RNAi line (from C. Rushlow lab, Sun et al., 2015). We used the MTD-GAL4 line alone as the control line.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
We performed ChIP-seq as previously described (Blythe and Wieschaus, 2015). We collected and fixed 200-400 embryos from each MTD-GAL4 and RNAi cross 0-2hr or 2-4hr after fertilization. We used 3 ul of rabbit anti-CLAMP (Soruco et al., 2013) and 2 ul rat anti-ZLD (from C. Rushlow lab) per sample. We performed three biological ChIP replicates for each protein (n=2), genotype (n=3) and time point (n=2). In total, we prepared 36 libraries using the NEBNext Ultra ChIP-seq kit (New England Biolabs) and sequenced libraries on Illumina HiSeq 2500 sequencer in 2x150-bp mode.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
CLAMP_MTD_0-2hr_avg_SES.bw
CLAMP_MTD_0-2hr_C1_x2value_bl.bed
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| Data processing |
We trimmed ChIP sequencing raw reads with Trim galore (v. 0.5.0, Krueger, 2017) with the minimal phred score 20, 36bp minimal read length and illumina adaptor removal. We then mapped cleaned reads to the D. melanogaster genome (UCSC dm6) with Bowtie2 (v. 2.3.0) with --very-sensitive-local flag. We used Picard tools (v. 2.9.2) and SAMtools (v.1.9, (Li et al., 2009) to remove the PCR duplicates. We used MACS2 (version 2.1.1, Zhang et al., 2008) to identify peaks with default parameters and MSPC (v.4.0.0, Jalili et al., 2015) to obtain consensus peaks from 3 replicates. ENCODE blacklist was used to filter out problematic regions in dm6 (Amemiya et al., 2019). We identified differential binding and non-differential binding using DiffBind with the DESeq2 method (v. 3.10, Stark and Brown, 2019). We used DeepTools (version 3.1.0, RamÃrez et al., 2014) and Homer (v 4.11, Givler and Lilienthal, 2005) to generate enrichment heatmaps (SES normalization), average profiles, motif searches, peak overlaps and peak annotation. Visualizations and statistical tests were conducted in R (R Core Team, 2014).
Genome_build: dm6
Supplementary_files_format_and_content: Peak files: consensus peaks from 3 replicates, obtained using MSPC (v.4.0.0). Peak files are in bed format. The column 5 has X^2 of the p-values of all the peaks overlapping this peak (including the peak itself) with 2k degree of freedom, where k is the number of overlapping peaks. Bigwig files: generated using Deeptools (v.3.1.0) bamCompare tools. Each IP and coresponding input file were compared in bamCompare, -SES option was used to normalize the library size difference. Log2 ratio of (IP/input) Fold Change value was ouput from bamCompare in bigwig format.
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| Submission date |
Jun 16, 2020 |
| Last update date |
Aug 05, 2021 |
| Contact name |
Erica Larschan |
| E-mail(s) |
erica_larschan@brown.edu
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| Organization name |
Brown University
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| Department |
MCB
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| Lab |
Larschan Lab
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| Street address |
185 meeting street, Room 368
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| City |
Providence |
| State/province |
Rhode island |
| ZIP/Postal code |
02912 |
| Country |
USA |
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| Platform ID |
GPL17275 |
| Series (2) |
| GSE152598 |
CLAMP and Zelda function together as pioneer transcription factors to promote Drosophila zygotic genome activation [ChIP-Seq] |
| GSE152613 |
CLAMP and Zelda function together as pioneer transcription factors to promote Drosophila zygotic genome activation |
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| Relations |
| BioSample |
SAMN15249978 |
| SRA |
SRX8556348 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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